GSM2433334: Whole WAT H3K27ac rep1; Mus musculus; ChIP-Seq
Sample information curated by ChIP-Atlas
Antigen
Antigen Class
Histone
Antigen
H3K27ac
Cell type
Cell type Class
Adipocyte
Cell type
White adipocytes
NA
NA
Attributes by original data submitter
Sample
source_name
epididymal WAT
strain
C57BL/6J
tissue
adipose
chip antibody
H3K27ac (Active Motif, 39133)
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Tissues were dounce homogenized, and cross-linked nuclei were isolated and sorted by BD FACS Aria II using mCherry fluorescence. Sorted nuclei were sheared by Covaris E220 and used for ChIP for overnight. Immunoprecipitates were washed and subjected to elution and reverse crosslinking. DNA was then extracted by AMPure XP beads according to the manufacturer’s manual. Extracted DNA (1-10ng, or all if less) was used to generate sequencing libraries by following the “on-bead” sequencing library preparation method. Briefly, DNA was processed through end repair/phosphorylation using the End-It DNA End-Repair Kit (Epicentre), A-tailing using the Klenow Fragment (NEB M0212) and index adaptor ligation using the Quick Ligase (NEB M2200). AMPure XP beads were left in all the reactions to clean up the DNA using PEG(Polyethylene Glycol 8000)/NaCl solution. After ligation, DNA was eluted from AMPure XP beads and then PCR-amplified using the PfuUltra II Hotstart PCR Master Mix (Agilent 600850). Gel electrophoresis/extraction was performed using the E-Gel EX Agarose Gels (Invitrogen) and MinElute Gel Extraction (Qiagen) to select library fragments between 250 and 600bp. Quantity and quality of the libraries were analyzed by Qubit and Agilent Bioanalyzer, respectively, and the libraries were pooled at a final concentration of 12pM and sequenced by HiSeq2500 or NextSeq 500 systems.